Spontaneous autoimmune myocarditis and cardiomyopathy in HLA-DQ8.NODAbo transgenic mice

Abstract

Most individuals have viral infections at some point in their life, however, only few develop autoreactivity to cardiac myosin following infection suggesting a genetic predisposition. Population studies have shown that among all the genetic factors linked with autoimmune disease development, MHC class II genes are the most significant genetic factors. Experimental autoimmune myocarditis resembling human Dilated cardiomyopathy can be induced in susceptible mice by infection with coxsackie virus as well as immunization with purified foreign and murine cardiac specific a-myosin. We generated transgenic mice lacking endogenous class II molecules, HLA-DR3.Abo and HLA-DQ8.Abo transgenic mice in NOD and HLA-DQ8.Abo in B10 background, to study the role of MHC in spontaneous autoimmunity. The HLA molecules in these mice are expressed on cell surface and can positively select CD4+ T cells expressing various Vb T cell receptors. NOD.DQ8 female mice spontaneously developed myocarditis and dilated cardiomyopathy. Histopathology of heart revealed mononuclear infiltrate consisting of CD4 and Mac-1+ cells and myocyte necrosis. NOD.DQ8 mice showed cellular and humoral autoreactive response to self cardiac myosin.. Depletion of CD8 and CD4 + cells suggested that CD8 T cells may act as regulatory cells while CD4 cells are required as effector cells. NOD.DR3 and B10.DQ8 mice did not develop any cardiac pathology suggesting DQ8 is required for predisposition to the spontaneous autoreactivity while NOD background influences onset and progression of disease. Thus these mice provide powerful tools to understand the role of HLA class II molecules in predisposition and onset of human diseases and to develop immunotherapy.

Figure 1

NOD.DQ8.Abo mice develop spontaneous myocarditis leading to dilated cardiomyopathy. A) Histopathology of heart shows mononuclear infiltration along with myocyte destruction and B) IgG deposition in heart. Hematoxylin and Eosin stained section of C) thyroid gland D) Liver E) Kidney and F) Pancreas do not show any histopathology. Micrographs are at X100 magnification.

Figure 2

A) Anti-tag antibodies in male and female NOD.DQ8.Abo mice show higher levels in female mice. However, no enteropathy was observed in these mice as observed by H&E staining of sections of Ileum. B) Peptide ELISA using overlapping peptides of tissue transglutaminase shows a higher reactivity of NOD.DQ8 mice compared to B10.DQ8 mice. The region comprising aa 100–130 contained the most immunogenic epitopes.

Figure 3

Echocardiographic assessment of heart of NOD.DQ8 and B10.DQ8 mice. A) Parasternal long axis view and B) Parasternal short axis view show dilation of both right and left inter-ventricle and left atrium with thickening of septum in hearts from NOD.DQ8 mice compared to B10.DQ8. RV-right ventricle, LV- left ventricle, LA-left atrium, and IV- intra ventricular.

Figure 4

Protocol for depleting CD4 and CD8 T cells in vivo. Mice were given intravenous injections of anti-CD4 (GK1.5) and anti-CD8 (lyt2) antibodies on indicated days. Presence of CD4 and CD8 cells in peripheral blood was analyzed by FACS after staining with specific conjugated antibodies on indicated days (top line). Most of the mice showed depletion of CD4 cells by 99% while CD8 T cells were depleted ranging between 85–95% in mice. Depleted mice were monitored for development of Myocarditis/dilated cardiomyopathy. CD4 depleted mice showed a later mortality as compared to nondepleted mice while CD8 depleted mice showed myocarditis in 100% of mice.

Figure 5

Spontaneous cytokine production in response to in vitro challenge with porcine cardiac myosin in male and female mice with cardiomyopathy and healthy NOD.DQ8.Aβo mice.

Figure 6

Schematic diagram of pathogenesis of myocarditis/dilated cardiomyopathy. Any damage to heart by viral infection can lead to cell death and availability of self antigen. Myocytes express class II molecules and in the event of inflammation, can present self antigen to infiltrating T cells. Alternatively, T cells and DCs in the periphery amplify the autoreactive response via antigen presentation, self or microbial epitopes that mimic heart antigen, and production of autoantibodies. These effector T cells can home to heart and lead to damage. Monocytes can carry antigens to heart and can activate DCs locally resulting in production of chemokines and inflammatory cytokines leading to infiltration of heart with T cells and mature DCs followed by destruction of myocytes and cardiomyopathy.